Inflammation represents a key event of many diseases, such as psoriasis, inflammatory bowel diseases, rheumatoid arthritis, asthma, multiple sclerosis, atherosclerosis, cystic fibrosis, and sepsis. Inflammatory cells, such as neutrophils, eosinophils, basophils, mast cells, macrophages, endothelial cells, and platelets, respond to inflammatory stimuli and foreign substances by producing bioactive mediators. These mediators act as autocrines and paracrines by interacting with many cell types to promote the inflammatory response. There are many mediators that can promote inflammation, such as cytokines and their receptors, adhesion molecules and their receptors, antigens involved in lymphocyte activation, and IgE and its receptors.
Cytokines, for example, are soluble proteins that allow for communication between cells and the external environment. The term cytokines includes a wide range of proteins, such as lymphokines, monokines, interleukins, colony stimulating factors, interferons, tumor necrosis factors, and chemokines. Cytokines serve many functions, including controlling cell growth, migration, development, and differentiation, and mediating and regulating immunity, inflammation, and hematopoiesis. Even within a given function, cytokines can have diverse roles. For example, in the context of mediating and regulating inflammation, some cytokines inhibit the inflammatory response (anti-inflammatory cytokines), others promote the inflammatory response (pro-inflammatory cytokines). And certain cytokines fall into both categories, i.e., can inhibit or promote inflammation, depending on the situation. The targeting of proinflammatory cytokines to suppress their natural function, such as with antibodies, is a well-established strategy for treating various inflammatory diseases.
Many inflammatory diseases are treated by targeting proinflammatory cytokines with antibodies. Most (if not all) of the anti-proinflammatory cytokine antibodies currently on the market, and those currently in clinical trials, are of the IgG class. See, for example, Nature Reviews, vol. 10, pp. 301-316 (2010); Nature Medicine, vol. 18, pp. 736-749 (2012); Nature Biotechnology, vol. 30, pp. 475-477 (2012); Anti-Inflammatory & Anti-Allergy Agents in Medicinal Chemistry, vol. 8, pp. 51-71 (2009); F1000.com/Reports/Biology/content/1/70, F1000 Biology Reports, 1:70 (2009); mAbs 4:1, pp. 1-3 (2012); mAbs 3:1, pp. 76-99 (2011); clinicaltrials.gov (generally), and clinicaltrialsregister.eu/ (generally). These IgG antibodies are administered systemically and thus are often associated with unwanted side effects, which can include one or more of, for example, infusion reactions and immunogenicity, hypersensitivity reactions, immunosuppression and infections, heart problems, liver problems, and others. Additionally the suppression of the target cytokines at non-diseased parts of the body can lead to unwanted effects.
In an attempt to reduce side effects associated with systemic treatment and to eliminate the inconvenience and expense of infusions, an article proposed an oral anti-TNF therapy that could be useful in treating Crohn's disease. Worledge et al. “Oral Administration of Avian Tumor Necrosis Factor Antibodies Effectively Treats Experimental Colitis in Rats.” Digestive Diseases and Sciences 45(12); 2298-2305 (December 2000). This article describes immunizing hens with recombinant human TNF and an adjuvant, fractionating polyclonal yolk antibody (IgY, which in chickens is the functional equivalent to IgG), and administering the unformulated polyclonal IgY (diluted in a carbonate buffer to minimize IgY acid hydrolysis in the stomach) to rats in an experimental rodent model of colitis. The rats were treated with 600 mg/kg/day of the polyclonal IgY. The uses of animal antibodies and polyclonal antibodies, however, are undesirable.
In a similar attempt to avoid adverse events associated with systemic administration, another group, Avaxia Biologics Inc., describes a topical (e.g., oral or rectal) animal-derived polyclonal anti-TNF composition that could be useful in treating inflammation of the digestive tract, such as inflammatory bowel disease. WO2011047328. The application generally states that preferably the polyclonal antibody composition is prepared by immunizing an animal with a target antigen, and the preferably the polyclonal antibody composition is derived from milk or colostrum with bovine colostrums being preferred (e.g., p. 14). The application also generally states that the animal derived polyclonal antibodies could be specific for (among other targets) other inflammatory cytokines (e.g., pp. 6-7). This application describes working examples in which cows were immunized with murine TNF and the colostrum was collected post-parturition to generate bovine polyclonal anti-TNF antibodies (designated as AVX-470). The uses of animal-derived antibodies and polyclonal antibodies, however, are undesirable.
IgA molecular forms have been proposed as treatments for various diseases, most notably as treatments for pollen allergies, as treatments against pathogens, and as treatments for cancer.
For example, one article describes anti-AmbαI (a ragweed pollen antigen) humanized monomeric IgA and dimeric IgA antibodies made in murine cells (NS0 and Sp2/0 cells). The dimeric IgA contains a mouse J-chain. The article proposes that the antibodies may be applied to a mucosal surface or the lower airway to inhibit entry of allergenic molecules across the mucosal epithelium and therefore to prevent the development of allergic response. Sun et al. “Human IgA Monoclonal Antibodies Specific for a Major Ragweed Pollen Antigen.” Nature Biotechnology 13, 779-786 (1995).
Several other articles propose the use of IgA antibodies as a defense against pathogens.
Two articles proposed the use of an anti-streptococcal antigen I/II secretory IgA-G hybrid antibody. Ma et al. “Generation and Assembly of Secretory Antibodies in Plants.” Science 268(5211), 716-719 (May 1995); Ma et al. “Characterization of a Recombinant Plant Monoclonal Secretory Antibody and Preventive Immunotherapy in Humans.” Nature Medicine 4(5); 601-606 (May 1998). The hybrid antibody contains murine monoclonal kappa light chain, hybrid Ig A-G heavy chain, murine J-Chain, and rabbit secretory component. The antibody was made by successive sexual crossing between four transgenic N. tabacum plants and filial recombinants to form plant cells that expressed all four protein chains simultaneously. The parent antibody (the source of the antigen binding regions, is identified as the IgG antibody Guy's 13. The group proposes that although sIgA may provide an advantage over IgG in the mucosal environment, such is not always the case (1998 Ma at p. 604, right column).
A related article identifies the anti-streptococcal antigen I/II secretory IgA-G hybrid antibody, which was derived from Guy's 13 IgA, as CaroRx. Wycoff. “Secretory IgA Antibodies from Plants.” Current Pharmaceutical Design 10(00); 1-9 (2004). Planet Biotechnology Inc. This related article states that the CaroRx antibody was designed to block adherence to teeth of the bacteria that causes cavities. Apparently, the CaroRx antibody was difficult to purify; the affinity of Protein A for the murine Ig domain was too low and protein G was necessary for sufficient affinity chromatography. Furthermore, the article states that several other chromatographic media had shown little potential as purification steps for the hybrid sIgA-G from tobacco leaf extracts. The article also indicates that the authors were unable to control for human-like glycosylation in tobacco, but that such was not a problem because people are exposed to plant glycans every day in food without ill effect.
WO9949024, which lists Wycoff as an inventor, Planet Biotechnology Inc. as the applicant, describes the use of the variable regions of Guy's 13 to make a secretory antibody from tobacco. The application contains only two examples—the first a working example and the second a prophetic example. Working Example 1 describes the transient production of an anti-S. mutans SA I/III (variable region from Guy's 13) in tobacco. The tobacco plant was transformed using particle bombardment of tobacco leaf disks. Transgenic plants were then screened by Western blot “to identify individual transformants expressing assembled human sIgA” (p. 25). Prophetic Example 2 states that in a transformation system for Lemna gibba (a monocot), bombardment of surface-sterilized leaf tissue with DNA-coated particles “is much the same as with” tobacco (a dicot). The prophetic example also stops at screening by immunoblot analysis for antibody chains and assembled sIgA, and states that the inventors “expect to find fully assembled sIgA.”
Another article proposed the use of an anti-RSV glycoprotein F IgA antibodies (mIgA, dIgA, and sIgA). Berdoz et al. “In vitro Comparison of the Antigen-Binding and Stability Properties of the Various Molecular Forms of IgA antibodies Assembled and Produced in CHO Cells.” Proc. Natl. Acad. Sci. USA 96; 3029-3034 (March 1999). The sIgA antibody was made in CHO cells sequentially transfected with chimeric heavy and light chains, human J-Chain, and human secretory component, respectively. Single clones were generated to express the mIgA (clone 22), the dIgA (clone F), and the sIgA (clone 6) (p. 3031).
Still other articles proposed, for example: (1) anti-HSV mIgA made in maize (Karnoup et al. Glycobiology 15(10); 965-981 (May 2005)) (which states that at that time there had been little success in the application of IgA class antibodies to therapeutic use because of the difficulty in producing the dimeric form in mammalian cells at economic levels); (2) anti-C. difficile toxin A chimeric mouse-human monomeric and dimeric IgA made in CHO cells (Stubbe et al. Journal of Immunology 164; 1952-1960 (2000)); (3) anti-N. meningitidis chimeric IgA antibodies were produced in BHK cells cotransfected with human J-Chain and/or human secretory component (Vidarsson et al., Journal of Immunology 166; 6250-6256 (2001)); (4) anti-Pseudomonas aeruginosa O6 lipopolysaccharide chimeric mouse/human mIgA1 made in CHO cells (Preston et al. Infection and Immunity 66(9); 4137-4142 (September 1998)); (5) anti-Plasmodium mIgA made in CHO cells (Pleass et al. Blood 102(13); 4424-4429 (December 2003)) (which states that unlike their parental mouse IgG antibodies, the mIgA antibodies failed to protect against parasitic challenge in vivo); and (5) anti-Helicobacter pylori urease subunit A sIgA and dIgA (Berdoz et al. Molecular Immunology 41(10); 1013-1022 (August 2004)).
For a review article discussing passive and active protection against pathogens at mucosal surfaces, see Corthésy. “Recombinant Immunoglobulin A: Powerful Tools for Fundamental and Applied Research.” Trends in Biotechnology 20(2); 65-71 (February 2002).
Still other articles propose the use of IgA antibodies as a treatment for cancer.
For example, one article describes a Phase Ia trial of a muring anti-transferrin receptor IgA antibody (Brooks et al. “Phase Ia Trial of Murine Immunoglobulin A Antitransferrin Receptor Antibody 42/6.” Clinical Cancer Research 1(11); 1259-1265 (November 1995)). Another article describes a human anti-Ep-CAM mIgA made in BHK (baby hamster kidney) cells (Huls et al. “Antitumor Immune Effector Mechanisms Recruited by Phase Display-Derived Fully Human IgG1 and IgA1 Monoclonal Antibodies.” Cancer Research 59; 5778-5784 (November 1999)). Still another article describes an anti-HLA Class II chimeric mIgA antibody made in BHK cells (Dechant et al. “Chimeric IgA Antibodies Against HLA Class II Effectively Trigger Lymphoma Cell Killing.” Blood 100(13); 4574-4580 (December 2002)). Yet other articles describe anti-EGFR mIgA or dIgA antibodies made in CHO, including Dechant et al. “Effector Mechanisms of Recombinant IgA Antibodies Against Epidermal Growth Factor Receptor.” Journal of Immunology 179; 2936-2943 (2007), Beyer et al. “Serum-Free Production and Purification of Chimeric IgA Antibodies.” Journal of Immunology 346; 26-37 (2009) (stating that as of 2009, IgA antibodies have not been commercially explored for problems including lack of production and purification methods), and Lohse et al. “Recombinant Dimeric IgA Antibodies Against the Epidermal Growth Factor Receptor Mediate Effective Tumor Cell Killing” Journal of Immunology 186; 3770-3778 (February 2011).
For a review article on anti-cancer IgA antibodies, see Dechant et al. “IgA antibodies for Cancer Therapy.”Critical Reviews in Oncology/Hematology 39; 69-77 (2001); states that compared with infectious diseases, the role of IgA in cancer immunotherapy is even less investigated).
It is desirable to have alternative antibody treatments for inflammatory diseases that preferably avoid the disadvantages of current systemic and previously-proposed topical treatments of inflammation.